Team:Toulouse/Project/Spreading

From 2014.igem.org

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       <h2>Spreading</h2>
       <h2>Spreading</h2>
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       <p>How can SubtiTree respect the environment ? How do we keep control on Subtitree?</p>
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       <p>How can SubtiTree respect the environment? How do we keep control on Subtitree?</p>
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<p class="title2">Auxotrophy</p>
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<p class="title2">Using a glutamin auxotroph <i>B. subtilis</i> strain</p>
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<p class="texte">
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To make the bacterium dependant on the tree and to avoid its spreading in the environment, it should be preferable to use a strain of <i>B.subtilis</i> which is auxotroph to a particular amino acid. The bacterium should be unable to synthesize one essential amino acid, and should find it in its environment. The Glutamine could be a good example since it is wide-spread in the phloem sap. It is the amino acid which is present in highest concentration in the phloem sap. If our bacterium is unable to synthesize the Glutamine, it will be obliged to take it in its close environment, that is to say the phloem sap.  
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To make the bacterium dependant on the tree and to avoid its spreading in the environment, it should be preferable to use a strain of <i>B.subtilis</i> which is auxotroph to a particular amino acid. The bacterium should be unable to synthesize one essential amino acid, and should find it in its environment. The glutamine could be a good example since it is wide-spread in the phloem sap. It is the amino acid which is present in highest concentration in the phloem sap. If our bacterium is unable to synthesize the glutamine, it will be obliged to take it in its close environment, that is to say the phloem sap.  
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Thus, if the bacterium is in the sap, it can grow normally without any deficiency since it uses the Glutamine present in the sap ; but if it escapes from the tree and a fortiori from the sap, it will not be able to survive for a long time. Indeed, Glutamine is found in very low quantities in the ground. This system should guarantee that the bacterium develops only in the tree and not elsewhere in the surroundings of the tree.<br/>  
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Thus, if the bacterium is in the sap, it can grow normally without any deficiency since it uses the glutamine present in the sap ; but if it escapes from the tree and a fortiori from the sap, it will not be able to survive for a long time. Indeed, glutamine is found in very low quantities in the ground. This system should guarantee that the bacterium develops only in the tree and not elsewhere in the surroundings of the tree.<br/>  
Auxotroph <i>B.sutbilis</i> strains already exist and are indexed in databases as BGSC (Bacillus Genetic Stock Center), so it is easy to find.   
Auxotroph <i>B.sutbilis</i> strains already exist and are indexed in databases as BGSC (Bacillus Genetic Stock Center), so it is easy to find.   
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<p class="title2">Non-sporing</p>
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<p class="title2">Preventing sporulation of <i>B. subtilis</i></p>
<p class="texte">
<p class="texte">
In order to limit the spreading of our bacterium, we decided to limit its lifespan to only one season. The bacteria should be injected in spring, grow during the summer and finally should be inactivated in fall.<br\>  
In order to limit the spreading of our bacterium, we decided to limit its lifespan to only one season. The bacteria should be injected in spring, grow during the summer and finally should be inactivated in fall.<br\>  
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Bacillus subtilis is a sporing bacterium : sporulation enable the microorganism to handle very harsh conditions and to spread tree to tree. Indeed, a spore is a very resistant form that is adapted for unfavourable conditions and for dispersal.<br/>  
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<i>Bacillus subtilis</i> is a sporing bacterium : sporulation enable the microorganism to handle very harsh conditions and to spread tree to tree. Indeed, a spore is a very resistant form that is adapted for unfavorable conditions and for dispersal.<br/>  
To keep the control on the development of SubtiTree, our strain should therefore be non-sporing. Thus, after a season of treatment, the sape become less nutritious, the temperature is low and the engineered bacterium cannot survive the following winter.<br/>
To keep the control on the development of SubtiTree, our strain should therefore be non-sporing. Thus, after a season of treatment, the sape become less nutritious, the temperature is low and the engineered bacterium cannot survive the following winter.<br/>
In addition, deleting all the engineered bacterial community every year puts a brake on the evolution due to random mutation = to keep control on the genetic constructions  
In addition, deleting all the engineered bacterial community every year puts a brake on the evolution due to random mutation = to keep control on the genetic constructions  
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<p class="title2">Toxin-antitoxin system</p>
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<p class="title2">Using a toxin-antitoxin system</p>
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<p class="texte">
The goal of this module is to prevent horizontal transfer between bacteria. Indeed, it is necessary to avoid any exchange of genetic material between wild type organisms and optimized organisms : it could be dangerous because of mutations, and considering ethics, it seems to be essential to avoid the spreading of synthetic genes.<br/>
The goal of this module is to prevent horizontal transfer between bacteria. Indeed, it is necessary to avoid any exchange of genetic material between wild type organisms and optimized organisms : it could be dangerous because of mutations, and considering ethics, it seems to be essential to avoid the spreading of synthetic genes.<br/>
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<p class="title2">Integrative plasmid?</p>
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<p class="title2">Using integrative plasmids</p>
<p class="texte">
<p class="texte">
All our constructions should be carried by integrative plasmids (pS.., pS.., pS..). Consequently, our different genetic modules would be integrated in the bacterium genome. The integration in the genome is more stable as the constructions are less likely to be transferred to other microorganisms. In addition to that, the expression of our genetic modules would not be dependant on a selective pressure based on an antibiotic resistance (as we can not inject antibiotics in the tree), allowing a high level of transcription in planta.  
All our constructions should be carried by integrative plasmids (pS.., pS.., pS..). Consequently, our different genetic modules would be integrated in the bacterium genome. The integration in the genome is more stable as the constructions are less likely to be transferred to other microorganisms. In addition to that, the expression of our genetic modules would not be dependant on a selective pressure based on an antibiotic resistance (as we can not inject antibiotics in the tree), allowing a high level of transcription in planta.  

Revision as of 19:48, 8 October 2014